Device for optically determining microbial colony population on growth surface

ABSTRACT

A viewer-comparator device is described for optically determining the density of microbial colony growth in and on an inoculated nutrient layer. In the preferred construction the device comprises a base, a magnifying lens supported thereon, means connected to the base for supporting a nutrient layercoated surface, this surface being located in position for viewing thereof through the lens, means to enable direct visual comparison by an observer between the appearance of the nutrient layer-coated surface and reproductions of microbial growth at a plurality of representative intervals over a range of growth densities of clinical interest and lighting means depending from the base. The comparison enabling means is located adjacent the position to be occupied by the nutrient layer-coated surface for simultaneous viewing of both the comparison enabling means and the coated surface through the lens. The lighting means are so directed that light rays emanating therefrom cannot pass to the lens by reflection or by direct passage from lighting means to lens. With the lighting means so disposed and directed, the microbial colonies will scatter the light reaching them and appear bright in an essentially dark field (i.e. no bright background).

0 Elite States atent 1 1 1 1 3,712,74fi Berger-0n 51 Jan. 23, 1973 [54] DEVICE FOR OPTICALLY DETERMINING MICROBIAL COLONY ABSTRACT POPULATION ON GROWTH SURFACE A viewer-comparator device is described for optically [75] Inventor: John A. Be S h d determining the density of microbial colony growth in N,Y and on an inoculated nutrient layer. In the preferred construction the device comprises a base, a magnify- [73] Asslgnee: General Elecmc Company ing lens supported thereon, means connected to the [22] Fil d; J l 29,1971 base for supporting a nutrient layer-coated surface, this surface being located in position for viewing [21] Appl' 167365 thereof through the lens, means to enable direct visual comparison by an observer between the appearance of 52 us. (:1. ..356/256, 356/36, 356/201, the nutrient layer-mated Surface and reproductions 356/208, 356/244, 356/246 microbial growth at a plurality of representative inter- 511 1111.01. ..Cl2b 1/0o,c121 1/00 W a range gmwth densities climcal [58] Field of Search ..3S6/36, as, 102, 201, 203, and lighting dependF'g fmm 356/244 246 256 The comparison enablmg means 1s located ad acent the position to be occupied by the nutrient layer- [56] References Cited coated surface for simultaneous viewing of both the comparison enabling means and the coated surface UNITED STATE PATENTS through the lens. The lighting means are so directed that light rays emanating therefrom cannot pass to the s ll T "356/208 X lens by reflection or by direct passage from lighting 2:495:9l2 1/1950 :23: means to lens. With the lighting means so disposed Primary ExaminerRonald L. Wibert Assistant Examiner-F. L. Evans Attorney-Jerome C. Squillaro et a1.

and directed, the microbial colonies will scatter the light reaching them and appear bright in an essentially dark field (i.e. no bright background).

6 Claims, 3 Drawing Figures PAIENTEDJM 23 I975 SHEET 2 0F 2 IN l/E IV TORI JOHN A. ERGER N- 4/%V4 HIS ATTORNEY DEVICE FOR OPTICALLY DETERMINING MICROBIAL COLONY POPULATION ON GROWTH SURFACE BAC KGROUND OF THE INVENTION Improved devices are described in U. S. Patent application Ser. No. 130,254 Bergeron for combined .sampling and culturing to enable a measure of the microorganism content of a liquid sample. The aforementioned patent application is assigned to the assignee of the instant invention and was filed Apr. 1, 1971. The referred structure is a long, thin tube (e.g. a disposable sterile, transparent, pre-plugged serological pipette) having internal area thereof coated with a culture medium.

When such internally coated devices are used to determine the content of colony-producing microorganisms'(i.e. bacteria, yeast or molds) in a liquid sample, e.g. urine, it is important to be able to quickly, easily and accurately determine the colony population density resulting from inoculation with the sample.

SUMMARY OF THE INVENTION The above objective is satisfied by the viewer/comparator devices described herein for optically determining microbial colony population density on a transparent surface having an inoculated nutrient layer disposed thereon. In its simplest construction the device consists of a base, at least one light source, means for connecting this light source to the base, means connected to the base for positioning the trans parent surface (e.g. the pipette of the aforementioned Bergeron application) and means for enabling direct comparison by an observer between the appearance of the transparent surface and reproductions of microbial growth at a plurality of representative intervals over some given range of clinical interest. The comparisonenabling means is located to permit simultaneous viewing of the transparent surface and the comparisonenabling means.

BRIEF DESCRIPTION OF THE DRAWING The exact nature of this invention as well as objects and advantages thereof will be readily apparent from consideration of the following specification relating to the annexed drawing in which:

FIG. 1 is a side elevational view partially in section showing a magnifier-viewer-comparator constructed according to this invention and adapted to'receive and hold a long, thin tube (e.g. a transparent, pre-plugged serological pipette) for visual examination;

FIG. 2 is a section taken on line 2--2 of FIG. I and FIG. 3 is a side elevational view ofa viewer/comparator device constructed according to this invention.

DESCRIPTION OF THE PREFERRED EMBODIMENT The magnifier-viewer-comparator device is exemplary of a hand-held type unit constructed according to this invention and comprises magnifying lens 11 mounted in frame 12 supported upon base 13, comparator card 14 mounted in open U-shaped frame 16 affixed to base 13, U-shaped spring clamp 17 for releasably holding pipette 18 and a pair of light bulbs (e.g. GE-243 bulbs) 19, 21 mounted in sockets 22, 23,

respectively. The upright legs 16a, 16b and lower leg 16c of frame 16 are shown slotted to accommodate card 14. Spring clamp 17 and sockets 22, 23 are affixed to mounting block 24 depending from base 13.

Pipette 18 (more completely described in the aforementioned Bergeron application) is made of a transparent material, preferably a plastic. Plug 26 of cotton or similar fibrous material is disposed in passage 27 at 0 the mouthpiece end of pipette 18. The inner surface of passage 27 is covered by a layer 28 of a suitable solid (stiff gel capable of supporting itself) culture medium. Layer 28 will have been inoculated by contact with a liquid sample being checked for microorganism content by drawing some of the sample up into the nutrient-lined bore to a predetermined height below the upper limit of layer 28. The liquid is then permitted to drain therefrom and after the tip of the pipette has been plugged, the unit is ready for colony development in the vertical position at room temperature or for incubation in a thermostatically controlled oven (not shown). Incubation is usually carried out at a temperature of about 37C for a period of about 18-24 hours. After incubation (or standing at room temperature for the requisite period), the pipette 18 is ready for examination for microbial growth (e.g. colonies of bacteria) in device 10. Pipette 18 is forced into the U- shaped spring clamp 17 and thenslid endwise therein to a position adjacent comparator card 14 as shown in FIGS. 1 and 2. If desired, the lower leg 16c of frame 16 may be slotted whereby with card 14 removed, pipette 18 may be directly positioned in clamp 17.

In this position the pipette 18 is ready for magnified viewing along approximately a 2-inch length adjacent the junction between the inoculated and control portions of layer 28. The control portion of this surface is, of course, that area of layer 28 remaining above the level to which the liquid sample had been raised in the pipette during inoculation. The growth characteristics in this 2-inch length provide the most information for the quantitative determination of microorganism content. Thus, it is of interest not only to be able to determine size and number of colonies, but also to be able to view the growth characteristics of the aforementioned junction.

The appearance of the pipette 18 may be readily compared to the standard displays: a, b, c, d, e, f, which are reproductions of growth characteristics of the microorganisms of interest at decadic intervals over the range of clinical interest (e.g. l0 l0 cells/cc). Preferably comparator card 14 is slotted as shown behind the position to be taken by pipette 18. The observer looking through lens 11 secs pipette 18 and the standards at some magnification (e.g. 2X), pipette 18 being located at or close to the focal length of lens 11. In this manner, visual readouts for microorganism content may be readily obtained.

By way of illustration, urine specimens containing more than 10 viable bacteria/cc are generally considered to be from patients with an active infection of the urinary tract. At the lower concentration (up to and including 10 bacteria/cc) the pipette will appear substantially free of colonies. Similarly, at very high concentrations (10 l0 bacteria/cc) the barrel of the pipette may appear (to the unaided eye) to be free of colonies except that at the margin between the inoculated and control nutrient areas a clearly defined ring occurs. Seeing this ring, the observer will recognize that individual colonies are not visible, because the colonies are microscopic or because the nutrient has become completely overgrown indicating, thereby, a very high bacteria concentration. At the concentration in the 10 l bacteria/cc range proportional densities of colonies per unit length of pipette will be seen.

Lighting means 19, 21 are located below base 13 and are directed so as to provide maximal oblique illumination of the portion of pipette 18 being examined. Although a single source of light may be employed, it is preferred to use a pair of lamps directed so that the lighting patterns of the two lamps crosses along the axis of pipette 18.

The placement of lamps 19, 21 relative to pipette 18 is such that rays of light emanating from these lamps and reaching the surface of pipette l8 intersect therewith such that these rays are not reflected to the lens 11 (and thereby to the eye of the observer). Also, with lamps 19, 21 disposed below base 13 there is no direct passage of light to lens 11 and the system functions essentially as a dark field viewer with the colonies receiving and scattering light. It is this scattered light that reaches the eye of the observer via lens 11 making the colonies visible as spots brighter than the surrounding, less-illuminated background.

Thus, it is required that light from the light source must not pass directly to, or be reflected to, the eye of the viewer and the intensity of illumination (light flux) from the light source must be high enough that the light scattered by the colonies that reaches the eye of the observer is of greater intensity than the ambient light reflected from the surface of the pipette 18. Any position of one or more light sources meeting these criteria may be employed.

Device may be used for the optical determination of microbial colony population density in ordinary ambient light conditions and, in fact, the ambient light may serve to illuminate the standards on comparator card 14 with or without the aid of light from the light source(s).

The power required for lighting bulbs 19, 21 may be provided by batteries (not shown) or the bulbs may be connected to an AC/DC power source (not shown).

Although the utilization of device 10 for the viewing of pipettes has been used to illustrate this invention, nutrient-coated glass slides or nutrient-lined test tubes may also be viewed and compared in the same manner using suitable variations of clamping means 17. [n the case of pipettes and test tubes the angle of incidence a with the central axis thereof of all light rays reaching the sampling/culturing device should be greater than 45.

Also, advantageously the mounting of sockets 22 and 23 on mounting block 24 may be such as to permit adjustment of the angle a.

The viewer-comparator device 30 of FIG. 3 is a somewhat larger unit providing non-magnified visual readout. In its simplest version, it consists of base 31, light source 32 connected to base 31 via front wall 33, vertical support bracket 34, light shield 36 and comparator disc 37.

During readout, the nutrient-lined pipette 38 is supported so as to provide angle aa the central axis thereof having a value of greater than 45 for the light emanating from lamp 32 and impinging thereon. Cleat 39 locates the lower end of the pipette and the upper end rests against the horizontal member 41 of frame 34.

Comparator disc 37 is essentially the same as comparator card 14 although the standards may be separated further permitting location of a slot therebetween to accommodate pipette 38 between any two standards for ease of comparison. The standards, of course, need not vary in steps of 10X the previous density, but may be in other multiples such as in increments of 5X the previous density.

The distance, which light shield 36 extends forward, is such as to prevent the passage of light rays directly from light source 32 to the eye of the observer. Elevating means (thumb screw 42) is provided to enable adjustment of the position of pipette 38 (and device 30) relative to the observer. The position of light bulb 32 is also adjustable to provide the requisite angle a.

Device 30 is of particular value in viewing large numbers of pipettes in rapid sequence as in the laboratory of a hospital due to the ease of positioning the pipettes for readout Device 10 can be made as a foldable unit much as, for example, foldable viewers for examining 35 mm transparencies are made.

The comparator card may be a photograph, a transparency or similar reproduction. in the case of a transparency it may be advantageous to use a ground glass plate as a backing for the transparency.

What l claim as new and desire to secure by Letters Patent of the United States is:

1. A viewer/comparator device for optically determining microbial colony population density on a transparent surface having an inoculated nutrient layer thereon comprising in combination:

a. an opaque base,

b. at least one light source,

c. means for mounting said light source on one side of said base,

means connected to said base for positioning a transparent tubular member so that at least a portion of the length thereof is located on the opposite side of said base from said light source in which position light rays from said light source must pass directly to said portion to impinge thereon at an angle of incidence of greater than 45 with the central axis of said tubular member,

. means for enabling direct comparison by an observer between the appearance of said portion and reproductions of microbial growth at a plurality of representative intervals over a range of clinical interest, said comparison-enabling means being disposed adjacent the location available for the positioning of said portion and f. a magnifying lens supported on said opposite side of said base being spaced from said portion and disposed for simultaneous viewing of said portion and said comparison-enabling means, said light source being so directed that light rays passing therefrom to said opposite side of said base cannot pass directly to, or be reflected to, said lens.

2. The viewer/comparator device of claim 1 wherein a pair of light sources are employed, said light sources being directed so that the patterns of illumination of v 6 the two lamps cross each other at the portion of the tutwo spaced points. bular member being examined. 5. The viewer/comparator device of claim 1 wherein 3. The viewer/comparator device of claim I wherein the positioning means is a spring clamp. the attitude fthe light source is adjust bl 6. The viewer/comparator device of claim 1 wherein 4. The viewer/comparator device of claim 1 wherein 5 the compansonenablmg means is 3 P p the positioning means supports the tubular member at 

1. A viewer/comparator device for optically determining microbial colony population density on a transparent surface having an inoculated nutrient layer thereon comprising in combination: a. an opaque base, b. at least one light source, c. means for mounting said light source on one side of said base, d. means connected to said base for positioning a transparent tubular member so that at least a portion of the length thereof is located on the opposite side of said base from said light source in which position light rays from said light source must pass directly to said portion to impinge thereon at an angle of incidence of greater than 45* with the central axis of said tubular member, e. means for enabling direct comparison by an observer between the appearance of said portion and reproductions of microbial growth at a plurality of representative intervals over a range of clinical interest, said comparison-enabling means being disposed adjacent the location available for the positioning of said portion and f. a magnifying lens supported on said opposite side of said base being spaced from said portion and disposed for simultaneous viewing of said portion and said comparisonenabling means, said light source being so directed that light rays passing therefrom to said opposite side of said base cannot pass directly to, or be reflected to, said lens.
 2. The viewer/comparator device of claim 1 wherein a pair of light sources are employed, said light sources being directed so that the patterns of illumination of the two lamps cross each other at the portion of the tubular member being examined.
 3. The viewer/comparator device of claim 1 wherein the attitude of the light source is adjustable.
 4. The viewer/comparator device of claim 1 wherein the positioning means supports the tubular member at two spaced points.
 5. The viewer/comparator device of claim 1 wherein the positioning means is a spring clamp.
 6. The viewer/comparator device of claim 1 wherein the comparison-enabling means is a photograph. 